1. Field of the Invention
The present invention relates generally to the treatment of cancer. More particularly, it concerns novel compounds useful for chemotherapy, methods of synthesis of these compounds and methods of treatment employing these compounds. The novel compounds are substituted anthracyclines having a three ring system related to anthracyclines such as daunorubicin, idarubicin, epirubicin, and doxorubicin which are known to have antitumor activity.
2. Description of Related Art
Resistance of tumor cells to the killing effects of chemotherapy is one of the central problems in the management of cancer. It is now apparent that at diagnosis many human tumors already contain cancer cells that are resistant to standard chemotherapeutic agents. Spontaneous mutation toward drug resistance is estimated to occur in one of every 106 to 107 cancer cells. This mutation rate appears to be independent of any selective pressure from drug therapy, although radiation therapy and chemotherapy may give rise to additional mutations and contribute to tumor progression within cancer cell populations (Goldie et al., 1979; Goldie et al., 1984; Nowell, 1986). The cancer cell burden at diagnosis is therefore of paramount importance because even tumors as small as 1 cm (109 cells) could contain as many as 100 to 1,000 drug-resistant cells prior to the start of therapy.
Selective killing of only the tumor cells sensitive to the drugs leads to an overgrowth of tumor cells that are resistant to the chemotherapy. Mechanisms of drug resistance include decreased drug accumulation (particularly in multi-drug resistance), accelerated metabolism of the drug and other alterations of drug metabolism, and an increase in the ability of the cell to repair drug-induced damage (Curt et al., 1984; and Kolate, 1986). The cells that overgrow the tumor population not only are resistant to the agents used but also tend to be resistant to other drugs, many of which have dissimilar mechanisms of action. This phenomenon, called pleiotropic drug resistance or multi-drug resistance (MDR), may account for much of the drug resistance that occurs in previously treated cancer patients. The development of drug resistance is one of the major obstacles in the management of cancer. One of the traditional ways to attempt to circumvent this problem of drug resistance has been combination chemotherapy.
Combination drug therapy is the basis for most chemotherapy employed to treat breast, lung, and ovarian cancers as well as Hodgkin""s disease, non-Hodgkin""s lymphomas, acute leukemias, and carcinoma of the testes. Combination chemotherapy uses the differing mechanisms of action and cytotoxic potentials of multiple drugs.
Although combination chemotherapy has been successful in many cases, the need still exists for new anti-cancer drugs. These new drugs could be such that they are useful in conjunction with standard combination chemotherapy, or these new drugs could attack drug resistant tumors by having the ability to kill cells of multiple resistance phenotypes.
A drug that exhibits the ability to overcome multiple drug resistance could be employed as a chemotherapeutic agent either alone or in combination with other drugs. The potential advantages of using such a drug in combination with chemotherapy would be the need to employ fewer toxic compounds in the combination, cost savings, and a synergistic effect leading to a treatment regime involving fewer treatments.
The commonly used chemotherapeutic agents are classified by their mode of action, origin, or structure, although some drugs do not fit clearly into any single group. The categories include alkylating agents, anti-metabolites, antibiotics, alkaloids, and miscellaneous agents (including hormones). Agents in the different categories have different sites of action.
Antibiotics are biologic products of bacteria or fungi. They do not share a single mechanism of action. The anthracyclines daunorubicin and doxorubicin (DOX) are some of the more commonly used chemotherapeutic antibiotics. The anthracyclines achieve their cytotoxic effect by several mechanisms, including inhibition of topoisomerase II; intercalation between DNA strands, thereby interfering with DNA and RNA synthesis; production of free radicals that react with and damage intracellular proteins and nucleic acids; chelation of divalent cations; and reaction with cell membranes. The wide range of potential sites of action may account for the broad efficacy as well as the toxicity of the anthracyclines (Young et al., 1985).
The anthracycline antibiotics are produced by the fungus Streptomyces peuceitius var. caesius. Although they differ only slightly in chemical structure, daunorubicin has been used primarily in the acute leukemias, whereas doxorubicin displays broader activity against human neoplasms, including a variety of solid tumors. The clinical value of both agents is limited by an unusual cardiomyopathy, the occurrence of which is related to the total dose of the drug; it is often irreversible. In a search for agents with high antitumor activity but reduced cardiac toxicity, anthracycline derivatives and related compounds have been prepared. Several of these have shown promise in the early stages of clinical study, and some, like epirubicin and idarubicin, are used as drugs. Epirubicin outsells doxorubicin in Europe and Japan, but it is not sold in the U.S.
The anthracycline antibiotics have tetracycline ring structures with an unusual sugar, daunosamine, attached by glycosidic linkage. Cytotoxic agents of this class all have quinone and hydroquinone moieties on adjacent rings that permit them to function as electron-accepting and donating agents. Although there are marked differences in the clinical use of daunorubicin and doxorubicin, their chemical structures differ only by a single hydroxyl group on C14. The chemical structures of daunorubicin and doxorubicin are shown in FIG. 1.
Doxorubicin""s broad spectrum of activity against most hematological malignancies as well as carcinomas of the lung, breast, and ovary has made it a leading agent in the treatment of neoplastic disease (Arcamone, 1981; Lown, 1988; Priebe, 1995). Since the discovery of daunorubicin and doxorubicin (FIG. 1), the mechanistic details of the antitumor activity of anthracycline antibiotics have been actively investigated (Priebe, 1995a; Priebe, 1995b; Booser, 1994).
Unfortunately, concomitant with its antitumor activity, DOX can produce adverse systemic effects, including acute myelosuppression, cumulative cardiotoxicity, and gastrointestinal toxicity (Young et al., 1985). At the cellular level, in both cultured mammalian cells and primary tumor cells, DOX can select for multiple mechanisms of drug resistance that decrease its chemotherapeutic efficacy. These mechanisms include P-gp-mediated MDR and MPR-rediated MDR, characterized by the energy-dependent transport of drugs from the cell (Bradley et al., 1988), and resistance conferred by decreased topoisomerase II activity, resulting in the decreased anthracycline-induced DNA strand scission (Danks et al., 1987; Pommier et al, 1986; Moscow et al., 1988.
Among the potential avenues of circumvention of systemic toxicity and cellular drug resistance of the natural anthracyclines is the development of semi-synthetic anthracycline analogues which demonstrate greater tumor-specific toxicity and less susceptibility to various forms of resistance.
The present invention seeks to overcome drawbacks inherent in the prior art by providing compositions of agents that display increased cytotoxicity when compared with doxorubicin and can prevent and/or overcome multi-drug resistance and exhibit reduced cardiotoxicity. This invention involves novel compounds that have utility as antitumor and/or chemotherapeutic drugs, methods of synthesizing these compounds and methods of using these compounds to treat patients with cancer. The invention is generally based on the discovery that anthracycline derivatives that have ring groups or other groups attached to their sugar moiety have a surprisingly strong ability to kill tumor cells. In one aspect, the invention relates to compounds that can form an alkyl bond with DNA via formaldehyde mediated cross-linking.
To design novel anticancer drugs with improved targeting of DNA, the inventors have studied in depth how anthracyclines interact with DNA. First, their studies of the energetics of anthracyclines binding to DNA and analysis of the x-ray diffraction data of monomeric anthracyclines complexes with DNA oligonucleotides led to the rational design of DNA-bisintercalating drugs. Second, the inventors"" studies of formaldehyde-mediated crosslinking of anthracyclines with DNA, which demonstrated the regioselectivity and base specificity of that process, led us to prepare novel drugs designed to form covalent bonds with N2 guanine of DNA. In vitro evaluation identified DNA-crosslinking anthracyclines WP809 and WP836 as unusually potent cytotoxic agents.
New anthracyline-based agents designed to interact and crosslink with DNA have been synthesized. Some of these analogs contain unique three ring system which is relatively stable. Synthesized compounds displayed activity significantly higher than that of parental daunorubicin or doxorubicin. In brief, in vitro the compound WP836 derived from doxorubicin was 500- to 25,000-fold more potent than doxorubicin in test performed in several cell lines. Similarly, the increased activity was also noticed for analog WP809 obtained from daunorubicin. Other analogs were also designed and synthesized. Observed activity and high potency indicate that the primary mechanism of action of these analogs is different from doxorubicin and daunorubicin.
The substituted anthracyclines having a three ring system are exemplified by those anthracyclines found in FIGS. 3-9. These actions produced substituted anthracyclines having a three ring system which exhibit activity substantially different from the activities of doxorubicin or daunorubicin. These compounds are active against doxorubicin resistant tumors and/or are more cytotoxic than doxorubicin against sensitive tumors, and the mechanism probably relates to the sequence-governed, base-specific alkylation of DNA. Other substituted anthracyclines are exemplified in FIGS. 11, 12, 14, and 15.
In some specific embodiments, the substituted anthracycline compounds have the general formula: 
wherein: R1 denotes any suitable group or combination of groups to form a nucleic acid intercalator or binding compound, including but not limited to hydrogen (xe2x80x94H) group, a hydroxyl group (xe2x80x94OH), a methoxy group (xe2x80x94OCH3), an alkoxy group having 1-20 carbon atoms, an alkyl group having 1-20 carbon atoms, an aryl group having 1-20 carbon atoms, a fatty acyl group having the general structure xe2x80x94Oxe2x80x94CO(CH2)nCH3, wherein n=an integer from 1 to about 20, or a fatty acyl group having the general structure xe2x80x94Oxe2x80x94CO(CH2)1(CHxe2x95x90CH)m(CH2)nCH3, wherein 1 is an integer between 1 to 3, m is an integer between 1 and about 6, and n is an integer between 1 to about 9; each of R2 and R3 is, independently of the other, a hydrogen (xe2x80x94H) group, a hydroxyl group (xe2x80x94OH), a methoxy group (xe2x80x94OCH3) or a double bonded oxygen moiety; R4 is a hydrogen (xe2x80x94H) group, a hydroxyl group (xe2x80x94OH), a methoxy group (xe2x80x94OCH3) or a halide; each of Y1 and Y2 is, independently of the other, a hydrogen (xe2x80x94H) group; a hydroxyl group (xe2x80x94OH); a methoxy group (xe2x80x94OCH3); or a double bonded oxygen, sulphur, or nitrogen group; R5-R12 are, independently, xe2x80x94H, xe2x80x94OH, a halide, xe2x80x94OR13, xe2x80x94SH, xe2x80x94SR13, xe2x80x94NH2, xe2x80x94NHR13, xe2x80x94N(R13)2, and R can additionally be a saccharide, with the proviso that both of R6 and R7 or both of R5 and R8 or both of R5 and R11 or both of R6 and R12 are involved in forming a three ring structure or either of R5 and R6 is independently a mercapto-haloalkyl group or ether alkyl group containing easy leaving groups [halogen like iodine or sulfonate esters (xe2x80x94OSO2R) like mesyl or tosyl] or ether alkyl group containing aziridine, oxirane, thiirane, oxetane, thietane rings; and R13 is a hydrogen (xe2x80x94H) group, a hydroxyl group (xe2x80x94OH), a methoxy group (xe2x80x94OCH3), an alkoxy group having 1-20 carbon atoms, an alkyl group having 1-20 carbon atoms, an aryl group having 1-20 carbon atoms, a fatty acyl group having the general structure xe2x80x94Oxe2x80x94CO(CH2)nCH3, wherein n=an integer from 1 to about 20, or a fatty acyl group having the general structure xe2x80x94Oxe2x80x94CO(CH2)1(CHxe2x95x90CH)m(CH2)nCH3, wherein 1 is an integer between 1 to 3, m is an integer between 1 and about 6, and n is an integer between 1 to about 9.
Certain specific embodiments of the anthracyclines of the invention are shown in FIGS. 3-9 and FIGS. 12-15.
In the specific methods disclosed in this patent, iodobutyraldehyde is employed to form the three ring structure, as described in Example 1 and shown in FIG. 2. However, a variety of longer, shorter, or different halogenated aldehydes may be employed in the place of the iodobutyraldehyde in the general procedure to cause variations in the three ring structure. Exemplary three ring structures created by the use of iodobutyraldehyde are shown in FIG. 10. Such structures may be modified by varying the starting halogenated aldehydes to have additional atoms in their rings, different atoms in the place of the nitrogen and oxygens therein, and different side groups. Three ring structures formed in this manner may, in the broadest embodiments of the invention be attached to any suitable nucleic acid intercalating group or binding group that will bring the alkylating function of the three ring group into proximity with DNA, including, but not limited to the anthracyclines disclosed herein.
Other anthracycline-based DNA alkylators are also described herein and some examples are shown in FIGS. 12-15 and their method of synthesis is described herein.
The present application also comprises methods of preparing anthracyclines. In devising the synthetic schemes and compounds of the present invention, the inventors have created a variety of novel compounds. These compounds are described elsewhere in the specification and figures, and are given xe2x80x9cWPxe2x80x9d numbers. The structure of a compound designated with a xe2x80x9cWPxe2x80x9d number is ascertainable by reviewing the specification and figures. Exemplary specific compounds that are encompassed by the invention are WP809, WP836, WP846, WP851, WP840, and WP885.
The invention also considers methods of treating a patient with cancer, comprising administering to the patient a therapeutically effective amount of the contemplated substituted anthracycline compounds and therapeutic kits comprising, in suitable container means, a pharmaceutically acceptable composition comprising the contemplated substituted anthracycline compounds.